Islet beta-cell failure is central to the development of type 2 diabetes and is contributed to by both hyperglycemia and hyperlipidemia. The protein neprilysin, has been shown in non-islet tissues to be upregulated with the chronically elevated glucose and fat levels seen in type 2 diabetes. While the function of neprilysin in islets has not been investigated, our preliminary data show that it is synthesized and active in islets and may play a role in the modulation of beta-cell function in response to islet stressors. Thus, the overall aim of this proposal is to elucidate the role of neprilysin under conditions associated with impaired beta-cell function, namely increased fat and glucose. We hypothesize that in states of chronically elevated fat and glucose, neprilysin activity is upregulated thereby promoting beta-cell dysfunction. The following studies will be performed to investigate: 1. The role of neprilysin in the impairment of insulin secretion induced by chronically elevated fat. Firstly, neprilysin deficient (NEP-KO) mice will be fed a high fat diet for 12 weeks and insulin secretion will be measured in vivo. Our preliminary data indicate that NEP-KO mice are protected from high fat diet-induced reductions in glucose-stimulated insulin secretion. Secondly, islets from NEP-KO mice will be cultured with free fatty acids and insulin secretion will be examined in vitro. 2. The role of neprilysin in the impairment of insulin secretion induced by chronically elevated glucose. Our preliminary data suggest that neprilysin may be upregulated under high glucose conditions and thereby may mediate the induction of oxidative stress. Firstly, NEP-KO mice will receive a 48-hour glucose infusion to induce hyperglycemia, then insulin secretion will be measured in vivo. Secondly, islets from NEP-KO mice will be cultured in high glucose and insulin secretion will be examined in vitro. 3. The role of the renin-angiotensin system (RAS) in mediating neprilysin's effects under chronically elevated fat and glucose conditions. Since neprilysin is a component of the RAS, isolated islets will be used to determine whether neprilysin's activity in this pathway is responsible for impaired beta-cell function. Neprilysin inhibition in the beta-cells may have beneficial outcomes in type 2 diabetes and so our findings will have significant implications for the development of therapeutics to reduce/prevent beta-cell dysfunction.